Abstract
Autophagy is a vacuolar pathway for the regulated degradation and recycling of cellular components. Beclin1, a Bcl2-interacting protein, is a well-studied autophagy regulator. Homozygous loss of Beclin1 in mice leads to early embryonic lethality. However, the role of Beclin1 in regulating the pluripotency of embryonic stem cells and their differentiation remains poorly explored. To study this, we generated Beclin1-Knockout (KO) mouse embryonic stem cells (mESCs) using the CRISPR-Cas9 genome-editing tool. Interestingly, Beclin1-KO mESCs did not show any change in the expression of pluripotency marker genes. Beclin1-KO mESCs also displayed active autophagy, suggesting the presence of Beclin1-independent autophagy in mESCs. However, loss of Beclin1 resulted in compromised differentiation of mESCs in vitro and in vivo due to misregulated expression of transcription factors. Our results suggest that Beclin1 may play an autophagy-independent role in regulating the differentiation of mESCs.
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Acknowledgements
This work is supported by funds to S.B.-M. from the Department of Science and Technology (DST) under the Woman Scientist A program (SR/WOSA/LS-15/2016(G)), funds to D.S. from the Wellcome Trust – Department of Biotechnology (DBT), India, Alliance (Intermediate Fellowship IA/I/12/1/500507), and funds to D.P. from the Department of Science and Technology (DST) (DST/INSPIRE/04/2017/001707). We thank the animal facility at National Centre for Cell Science (NCCS), Pune. We thank Dr. Avinash Pradhan, King Edward Memorial (K.E.M.) Hospital, Pune, for his guidance on the histopathological examination of the teratomas. We thank members of D.S.’s lab for their inputs during the preparation of this manuscript.
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Corresponding editor: Kundan Sengupta
Corresponding editor: Kundan Sengupta
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Bivalkar-Mehla, S., Puri, D., Singh, S.B. et al. Understanding the role of Beclin1 in mouse embryonic stem cell differentiation through CRISPR-Cas9-mediated gene editing. J Biosci 46, 18 (2021). https://doi.org/10.1007/s12038-021-00139-5
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DOI: https://doi.org/10.1007/s12038-021-00139-5